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Molecular basis for Epstein-Barr virus genome DNA replication

EB 病毒基因组 DNA 复制的分子基础

基本信息

批准号：
12470073
负责人：
TSURUMI tatsuya
金额：
$ 5.38万
依托单位：
Division of Virology Aichi Cancer Center Research Institute
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2002
项目状态：
已结题

项目摘要

Productive infection/replication of herpesviruses usually occurs in growth arrested cells but there has been no direct evidence in case of Epstein-Barr virus (EBV) since an efficient lytic replication system without external stimuli does not exist for the virus. Expression of EBV lytic switch transactivator BZLF1 protein in EBV-negative epithelial tumor cell lines, however, is known to arrest cell cycle in G0/G1 by induction of the tumor suppressor protein, p53 and the cyclin dependent kinase inhibitors, p21WAF-1/CIP-1 and p27KIP-1 followed by the accumulation of hypophosphorylated form of Rb protein. In order to determine the effect of onset of the lytic viral replication on cellular events in EBV-latently infected B lymphoblastoid cell lines, a tightly controlled induction system of the EBV lytic replication program by inducible BZLF1 protein expression was established in B95-8 cells. The induction of the lytic replication completely arrested cell cycle progression and cellular DNA replication. Surprisingly, the levels of p53, p21WAF-1/CIP-1, and p27KIP-1 were constant before and after the lytic program induction, indicating that the cell cycle arrest induced by the lytic program is not mediated through p53 and the CDK inhibitors. Furthermore, although cellular DNA replication was blocked, elevation of cyclin E/A expression and accumulation of hyperphosphorylated forms of Rb protein were observed, being a post-G1/S phase characteristics of cells. Thus, while the EBV lytic program promoted specific cell cycle associated activities involved in the progression from G1 to S phase, it inhibited cellular DNA synthesis. Such cellular condition appears to most favors viral lytic replication.

疱疹病毒的生产性感染/复制通常发生在生长受阻的细胞中，但由于没有外部刺激的有效裂解复制系统，因此没有直接证据表明eb病毒（EBV）。然而，在EBV阴性上皮肿瘤细胞系中，EBV裂解开关反激活因子BZLF1蛋白的表达，已知通过诱导肿瘤抑制蛋白p53和周期蛋白依赖性激酶抑制剂p21WAF-1/CIP-1和p27KIP-1，然后积累低磷酸化形式的Rb蛋白，在G0/G1期阻滞细胞周期。为了确定裂解病毒开始复制对EBV潜伏感染的B淋巴母细胞样细胞系细胞事件的影响，在B95-8细胞中建立了一个通过诱导表达BZLF1蛋白来严格控制EBV裂解复制程序的诱导系统。裂解复制的诱导完全阻止了细胞周期的进展和细胞DNA的复制。令人惊讶的是，在裂解程序诱导前后，p53、p21WAF-1/CIP-1和p27KIP-1的水平是恒定的，这表明裂解程序诱导的细胞周期阻滞不是通过p53和CDK抑制剂介导的。此外，尽管细胞DNA复制被阻断，但观察到细胞周期蛋白E/A表达升高和Rb蛋白过度磷酸化形式的积累，这是细胞g1 /S期后的特征。因此，虽然EBV裂解程序促进了从G1期到S期的特定细胞周期相关活动，但它抑制了细胞DNA合成。这种细胞状态似乎最有利于病毒裂解复制。